1. Field of the Invention
This invention relates to the separation of particles from a liquid in which the particles are suspended, more particularly, the separation of blood cells from the blood plasma or the blood serum in which they are suspended.
2. Discussion of the Art
For in vitro diagnostics, biological samples currently used are samples of blood plasma or samples of blood serum. Disease markers related to proteins, lipoproteins, hormones, antibodies, antigens, virus, bacteria, parasites are commonly detected in blood plasma or blood serum of a patient. In order to collect blood plasma or blood serum, red blood cells, white blood cells, platelets, and other components must be removed from a sample of whole blood. Blood plasma makes up about 55% of total blood volume. It is composed mostly of water (90% by volume) and contains dissolved proteins, glucose, clotting factors, mineral ions, hormones, and carbon dioxide (plasma being the main medium for excretory product transportation). Blood serum is blood plasma without fibrinogen or the other clotting factors. Blood cells must be removed from blood plasma or blood serum before the sample of blood can be analyzed.
Centrifugation and filtration are currently used to separate blood cells from blood plasma or blood serum for diagnostic purposes. Both techniques require extensive labor and a relatively great amount of time for medical laboratories, which have limited resources with respect to both equipment and personnel. The drawbacks of centrifugation, wherein whole blood samples are introduced into a centrifuge rotating at 3000-3400 rpm for 10 to 15 minutes, include consumption of time, which results from the time needed by a technician to load and unload samples, the need for a skilled technician to aspirate blood plasma or blood serum with a pipette from the separated layers in blood collection tubes. The drawbacks of filtration processes include filter fouling and low throughput after fouling occurs. Other potential problems include breakage of blood collection tubes and loss of the sample. There is also the risk of hemolysis and the consequent destruction of the sample. Accordingly, it would be desirable to provide a method that is cost effective and efficient for the separation of blood cells from blood plasma or blood serum in order to analyze a sample of blood.
U.S. Patent Application Publication No. 2006/0021437 A1 discloses an apparatus and a method for concentrating analytes within a fluid flowing through a tube using acoustic radiation pressure. The apparatus includes a function generator that outputs a radio frequency electrical signal to a transducer that transforms the radio frequency electric signal to an acoustic signal and couples the acoustic signal to the tube. The acoustic signal is converted within the tube to acoustic pressure that concentrates the analytes within the fluid.
U.S. Pat. No. 5,711,888 discloses separation and recycling of particulate material suspended in a fluid by means of an ultrasonic resonance wave. In a preferred embodiment, the ultrasonic resonance field is generated within a multilayered composite resonator system including a transducer, the suspension, and a mirror parallel to each other. Dimensions and frequencies resonant to the whole system but not exciting Eigen-frequencies of transducer and mirror itself are chosen so that thermal dissipation is minimized. Specialized applications in biotechnology are described including an acoustic filter for mammalian cell bioreactors or the selective retention of viable cells relative to nonviable cells.
WO 2006/032703 A1 discloses a method and a device for separating particles using ultrasonic standing waves which are switched between two different frequencies. A second order harmonic standing wave is used together with a fundamental standing wave. If the particles are exposed to the fundamental standing wave, the forces act to collect particles at the center. If the particles are exposed to the second order harmonic standing wave, the forces act to collect particles at the two pressure nodes at the sides. By switching the frequency between the second order harmonic standing wave and the fundamental standing wave, particles with different properties will be exposed to different accelerations and are separated into two streams.
U.S. Pat. No. 3,832,655 discloses an ultrasonic delay line which comprises a solid body and two input and output electro-mechanical transducers for converting electrical energy into ultrasonic mechanical energy or vice versa, and in which the ultrasonic wave, emitted from the input electro-mechanical transducer by the application of an electrical input signal thereto, is reflected by at least one reflecting surface formed in the solid body and enters the output electro-mechanical transducer to derive therefrom an electric output signal which is delayed behind the electric input signal for a period of time during which the ultrasonic wave propagates in the solid body. The reflecting surface has at least one elliptical surface whose focuses are located each at one point on each electro-mechanical transducer or its equivalent point.
U.S. Pat. No. 4,055,491 discloses apparatus and method for using ultrasonic waves for removing microscopic particles from a liquid medium, such as algae from a solar or refuse pond, or blood cells from blood. The apparatus includes an ultrasonic generator propagating ultrasonic waves of over one megacycle per second through the liquid medium to cause the flocculation of the microscopic particles at spaced points. In two embodiments, the ultrasonic waves are propagated in the horizontal direction through the liquid medium, and baffle plates are disposed below the level of propagation of the ultrasonic waves. The baffles are oriented to provide a high resistance to the horizontal propagation therethrough of the ultrasonic waves and a low-resistance to the vertical settling therethrough of the flocculated particles. The ultrasonic generator is periodically energized to flocculate the particles, and then de-energized to permit the settling of the flocculated particles through the baffle plates from whence they are removed.
U.S. Pat. No. 4,673,512 discloses the separation of different types of particulate matter in a carrier liquid by using ultrasonic standing wave and relying on the different acoustic responses of the different particle types. By varying the acoustic energy propagation cyclically a more effective separation rate can be obtained, with a more readily attracted particle type being subjected to a further discrimination step in each cycle. The cyclical energy variation may be in the intensity of the standing wave, e.g., using suppression means, and/or the velocity of the standing wave relative to the liquid medium, e.g., using phase control means.
Pui et al., Batch and Semicontinuous Aggregation and Sedimentation of Hybridoma Cells by Acoustic Resonance Fields, Biotechnol. Prog. 1995, 11, 146-152, discloses the use of ultrasound to enhance the sedimentation of hybridoma cells from medium in a 75 mL resonator chamber. Forces in the acoustic standing waves aggregated the cells, and the aggregates were then rapidly sedimented by gravity. Cell separation increased with acoustic treatment time and cell concentration.
Gaida et al., Selective Retention of Viable Cells in Ultrasonic Resonance Field Devices. Biotechnol. Prog. 1996, 12, 73-76, discloses a double chamber ultrasonic resonance field device for the separation and retention of animal cells. By controlling operational parameters such as flow and power input, the device can retain viable cells more efficiently, allowing for selective removal of nonviable cells and cell debris.
Other techniques for separating blood cells from blood plasma or blood serum include electro-osmotic flow, which involves separation by size differential, which requires a conductive medium in a strong electric field, centrifugal force, as described in U.S. Pat. No. 5,186,844, magnetic separation, which requires the generation of a magnetic field either by high current or mechanical movement of magnets, dielectrophoretic separation, which requires high voltages with a non-conductive medium, as described in U.S. Pat. No. 6,881,314, electrophoretic separation, which requires high voltages with a conductive medium, such as electrolytes, as described in U.S. Pat. No. 6,881,314, diffusion-based separation, as described in U.S. Pat. No. 6,297,061, and optical trapping, which requires a single beam infrared laser, as described in U.S. Pat. No. 4,893,886.
Acoustic radiation forces can be expressed by the following equation:
            F      st        =                            2          ⁢                                    π              ⁡                              (                                  κ                  ⁢                                                                          ⁢                  R                                )                                      3                    ⁢          2          ⁢                      E            st                                    κ          2                    ⁢              Φ        ⁡                  (                      Λ            ,            σ                    )                    ⁢              sin        ⁡                  (                      2            ⁢            κ            ⁢                                                  ⁢                          r              0                                )                                Φ      ⁡              (                  Λ          ,          σ                )              =                  1        3            ⁢              (                                                            5                ⁢                Λ                            -              2                                                      2                ⁢                Λ                            +              1                                -                      1                          Λσ              2                                      )                            where        FS, represents the primary acoustic force acting on a particle;        Est represents the energy density of standing waves;        Λ represents the ratio of the density of the particle to the density of the fluid;        σ represents the ratio of the velocity of sound of the particle to the velocity of sound of the fluid;        R represents the radius of the particle;        ro represents the vector normal to the force node; and        k represents the sound wavenumber (sound frequency).See, for example, Kapishnikov et al., Continuous particle size separation and size sorting using ultrasound in a microchannel, Journal of Statistical Mechanics: Theory and Experiment, IOP Publishing, 2006, pages 1-15, incorporated herein by reference. The acoustic radiation force on a particle is influenced by the size, density, compressibility, and location of the particle, and the frequency and amplitude of acoustic radiation.        